Glutamine metabolism as potential target for prostate cancer radiosensitization

Background: A major reprogramming of cellular energy metabolism is a hallmark of tumor cells. In addition to an increased glucose uptake, highly proliferative cancer cells require additional supplies for their biosynthesis and energy production such as glutamine. Glutaminolysis also contributes to the ROS scavenging and activation of the pro-survival signaling pathways. Tumors with enhanced MYC expression, such as prostate cancer have a particularly high demand for glutamine. Herein, we investigated the role of glutamine metabolism pathways for prostate cancer radioresistance. Methods: Prostate cancer cell lines DU145, PC3, LNCaP and their radioresistant sublines (RR) were analyzed by metabolomic and gene expression profiling. The relative cell sensitivity to the inhibition of glutaminolysis was measured by analysis of viability (MTT), apoptosis and necrosis (flow cytometry and Western blotting), levels of ROS and glutathione (flow cytometry), radiosensitivity (colony formation assay, CFA), DNA repair (γH2A.X foci) and tumorigenicity in mice. Primary cell cultures from 12 tumor biopsies and matched benign tissues from prostates cancer patients were characterized by radiobiological 3D CFA and by gene expression profiling, and relative radioresistance was correlated with expression levels of the genes regulating glutaminolysis. The Cancer Genome Atlas (TCGA) datasets were analyzed for correlation of the gene expression levels and patients outcome. Results: Glutaminolysis is upregulated in RR cells, where glutamine is mostly used for production of α-ketoglutarate, which is involved in ROS scavenging and epigenetic resetting by regulation of the histone methylation, whereas α-ketoglutarate utilization for Krebs cycle is suppressed. Deprivation of glutamine or siRNA mediated inhibition of glutaminolysis leads to the induction of endoplasmic reticulum (ER) stress and inhibition of the DNA repair, clonogenicity and in vivo tumorigenicity after irradiation with a more pronounced effect for RR cells. Analysis of the TCGA datasets revealed that a high expression of the genes regulating prostate cancer glutaminolysis is significantly associated with a decrease in relapse free survival after radiotherapy. Discussion: Prostate cancer cell radioresistance is associated with alterations of glutaminolysis, whose inhibition increases the cytotoxic effects of radiation in prostate tumor cells. Expression of the proteins involved in glutaminolysis can be potentially used to predict clinical outcome after radiation therapy.